Lysophosphatidic acid (LPA) is a major mitogen in serum that regulates an array of cellular processes related to pathogenesis of cancer and other human diseases. Despite the central role of LPA in controlling cell growth and other cellular activities, very little is understood about the signaling mechanisms of LPA. While protein tyrosine phosphorylation has been recognized as an important signaling mechanism of LPA and other agonists of G protein-coupled receptors, it remains largely unknown how activation of G-proteins leads to tyrosine phosphorylation. To address this important question, LPA-induced tyrosine phosphorylation of the epidermal growth factor receptor (EGFR) will be studied. Based on our preliminary observations and increasing evidence of redox-regulation in cell signaling, it is postulated that LPA transactivates the EGFR by decreasing EGF receptor phosphatase activity through a mechanism mediated by calcium and reactive oxygen species (ROS). Three specific aims are proposed to critically evaluate key aspects of this hypothesis. In Specific Aim I, the requirement for intrinsic EGFR tyrosine kinase activity will be assessed using kinase- defective EGFR and new specific inhibitors for the EGFR. Effects of decreasing protein tyrosine phosphatase activity toward EGFR will also be examined. Alternatively, a novel sensitive approach is proposed to determine whether LPA activates the EGFR kinase. TO further evaluate whether LPA-induced tyrosine phosphorylation is catalyzed by intrinsic EGF receptor kinase or by another cellular kinase, Specific Aim II proposes phosphopeptide mapping and mutagenesis experiments to analyze LPA-induced tyrosine phosphorylation sites on the EGFR. In Specific Aim III, regulation of EGFR-dephosphorylating activity by LPA will be investigated both in vitro and in intact cells under conditions that redox-mediated changes can be detected, and the involvement of calcium and ROS will be evaluated. These studies not only will advance our understanding of signal transduction by LPA and G-proteins that control many cellular processes fundamental to the development of cancer and other human diseases, but also will stimulate further research in the emerging area of oxidative signaling.